Bootcongres

Aim: Plasmacytoid dendritic cells (pDC) are important mediators of anti-viral immune responses, during which they produce vast amounts of interferon (IFN)-α. In recent years pDC have also been characterized as antigen presenting cells (APC), capable of phagocytosing both soluble and cellular antigen, and to subsequently activate CD4⁺ and CD8⁺ T cells. Therefore these cells appear to be at the crossroads of allo- and anti-viral-immunity. We previously demonstrated a strong influx of pDC in the tubulointerstitium of renal biopsies with acute rejection, however the exact role of pDC in the process of rejection remains elusive. Methods: pDC were isolated from buffy coats of healthy volunteers by negative selection, and were cocultured with either viable or necrotic human renal epithelial HK2 cells. The cocultures were performed in the absence or presence of the viral DNA mimic CpG, and pDC were analysed for the expression of costimulatory molecules (FACS) and cytokine production (ELISA). Results: We show that activation of pDC using CpG, led to induction of IFN-α production and upregulation of both co-stimulatory (CD83, CD86) and co-inhibitory (PD-L1) molecules, whereas the pDC lineage markers BDCA-2, CD123, and CD11c were unaffected. Co-culture of pDC with either viable or necrotic HK2 cells did not affect the expression of these markers. Using CFSE labelled HK2, we could demonstrate that necrotic material was ingested by pDC, provided that they were activated with CpG. Activation by CpG in the presence of necrotic HK2 cells did not significantly affect pDC marker expression, compared to activation with CpG alone. To mimic actual viral infections, we are currently exploring infection of HK2 cells with a GFP-labelled cytomegalovirus strain, and optimized the infection of HK2 cells reaching an efficiency of 50% infected cells. 

Conclusion: Our data suggest that although pDC are strongly activated by viral mimics, the modulating effects of either viable or dying HK2 renal cells appears limited. However, this should be further explored in the context of actual virus-infected renal cells, and for the full APC function of pDC, including T cell stimulatory capacity.